The development of the mammalian immune system is typically thought to occur in a linear fashion, from immaturity to maturity as a function of antigen exposure. Previous findings in birds and in mice, however, indicate that this view is oversimplified. Thus, in these species, the developing immune system appears to be layered in a manner that is independent of antigen exposure, beginning as a multilineage fetal system that is replaced by an anatomically and biologically distinct multilineage system after birth. If so, then developmentally ordered and unique hematopoietic stem/progenitor cells (HSPC) could give rise to distinct lymphocyte lineages at different stages of development. In ongoing experiments, we have found that such immune system layering occurs in humans. Our preliminary data show that a vigorous human fetal immune response to exogenous antigens can be actively suppressed by antigen-specific Tregs, that these fetal Tregs are derived from a fetal-specific lineage of T cells, and that this lineage is generated by an HSPC that is distinct from that found in adults. These data suggest that the human immune system is comprised of two distinct waves: one generated from a fetal HSPC that exists in utero in the fetal liver and bone marrow, and another generated from a superseding adult HSPC that resides in the bone marrow at later time points. The former gives rise to an immune system that is prone to deliver a tolerogenic response to foreign antigens. The latter gives rise to an immune system that is more likely to generate an immunoreactive response (e.g., one including cytotoxic T cells and neutralizing antibodies). Given these findings, we hypothesize that physiologic layering of immune system ontogeny leads to a normal range in the ratio of fetal- to adult-type T cells at birth, with some neonates exhibiting a higher fraction of fetal T cells than others; and that those with a high ratio of fetal/adult T cells will generate predominant Th2 responses to routine childhood immunizations. These hypotheses will be addressed in the experiments of the following Specific Aims: (1) to determine the normal range of fetal to adult T cells in the umbilical cord blood of the full term neonate; and (2) to determine whether those full term neonates with a high ratio of fetal/adult T cells are more likely to generate a Th2-polarized immune response to routine childhood vaccines. Should this exploratory study reveal normal variation in the ratio of fetal to adult T cells at birth and should such variation be directly related to a Th2 skew after childhood vaccination, modalities aimed at changing this ratio more towards the adult lineage at birth may provide benefit to a substantial number of newborns. PUBLIC HEALTH RELEVANCE: These exploratory studies are relevant to public health for two reasons. First, they may provide proof-of- concept evidence for the existence of a range in the extent of immune system layering in human neonates. Secondly, they may demonstrate that this range is itself related to (and possibly causal of) differences in the ability of neonates to withstand infections and to respond to vaccines.